​​Brain processes underlying behavior, cognition, and emotion involve communication between neurons by the regulated release of neurotransmitter from synaptic vesicles. Synaptic vesicles reside in several functional pools in the presynaptic terminal that are defined by their location and probability of fusion upon stimulation. The high frequency of transmitter release observed at many synapses requires mechanisms to recycle synaptic vesicle membrane, proteins, and transmitter locally at the nerve terminal. Variation in the kinetics of vesicle recycling or mobilization from vesicle pools may shape the amount and pattern of neurotransmitter output, and hence contribute to information processing and some forms of synaptic plasticity. We are using a variety of complementary approaches‚Äîincluding biochemistry, live cell imaging, and mouse genetics‚Äîto understand how trafficking of synaptic vesicle components regulates synaptic transmission.

Multiple mechanisms have been proposed to mediate the recycling of synaptic vesicles, but most models assume that the protein components of the vesicles recycle together. However, specific sorting signals and protein interactions of the vesicular glutamate transporter, VGLUT1, direct recycling of this protein to different pathways. Comparison of VGLUT1 to other synaptic vesicle proteins reveals kinetic and mechanistic differences, indicating that the recyclingof other proteinsmay be independently regulated. We are investigating the molecular mechanisms and regulation of individual synaptic vesicle protein recycling, which could result in activity-dependent alterations of synaptic vesicle (and plasma membrane) protein composition influencing transmitter release. These mechanisms may contribute to the different physiological properties observed at different types of synapses

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We demonstrated that VGLUT1 recycles in an activity-dependent manner to a novel pathway that uses the clathrin adaptor protein AP3 in addition to the well-described AP2 pathway. We are interested in the molecular mechanisms and functional consequences of targeting to different membrane trafficking pathways. The AP3 pathway may be particularly important in the recycling of synaptic vesicles from endosomes generated after prolonged stimulation, which may selectively replenish different vesicle pools. In addition, the endosomal pathway may be important in re-sorting proteins into functional synaptic vesicles or trafficking to a degradative pathway. Indeed, little is known about the lifetime and turnover of any synaptic vesicle protein. In the case of vesicular neurotransmitter transporters, protein levels could have significant effects on the amount of neurotransmitter stored and released.

We use and develop optical tools to image the dynamics of synaptic vesicle components. Fusions of vesicular neurotransmitter transporters with a pH-sensitive GFP will provide optical probes for imaging activity of neurotransmitter systems in individual neurons, circuits, and networks. These probes can be genetically encoded and targeted to specific neuronal populations, increasing the spatial resolution to study these systems. We are interested in studying the effect of drugs, treatments, or genes on neurotransmitter release in specific brain regions and circuits that may be relevant to neuropsychiatric disease. Our long term goal is to use these tools to gain insight into the pathophysiology of neuropsychiatric diseases and aid the development of novel therapeutics that alter synaptic transmission and behavior by altering neurotransmitter release.